Disruption of the interaction between mutationally activated Gq and G¦ attenuates aberrant signaling

Heterotrimeric G protein stimulation via G protein-coupled receptors promotes downstream proliferative signaling. Muta-tions can occur in G alpha proteins which prevent GTP hydrolysis; this allows the G proteins to signal independently of G protein- coupled receptors and can result in various cancers, such as uveal melanoma (UM). Most UM cases harbor Q209L, Q209P, or R183C mutations in G alpha q/11 proteins, rendering the proteins constitutively active (CA). Although it is generally thought that active, GTP-bound G alpha subunits are dissociated from and signal independently of Gfl gamma, accumulating evidence indicates that some CA G alpha mutants, such as G alpha q/11, retain binding to Gfl gamma, and this interaction is necessary for signaling. Here, we demonstrate that disrupting the interaction between Gfl gamma and G alpha q is sufficient to inhibit aberrant signaling driven by CA G alpha q. Introduction of the I25A point mutation in the N-terminal alpha helical domain of CA G alpha q to inhibit Gfl gamma binding, over-expression of the G protein G alpha o to sequester Gfl gamma, and siRNA depletion of Gfl subunits inhibited or abolished CA G alpha q signaling to the MAPK and YAP pathways. Moreover, in HEK 293 cells and in UM cell lines, we show that G alpha q-Q209P and G alpha q-R183C are more sensitive to the loss of Gfl gamma interaction than G alpha q-Q209L. Our study challenges the idea that CA G alpha q/11 signals independently of Gfl gamma and demonstrates differential sensitivity between the G alpha q-Q209L, G alpha q-Q209P, and G alpha q- R183C mutants.